Perkinsus marinus (Mackin et al.) Levine
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ATCC determines the biosafety level of a material based on our risk assessment as guided by the current edition of Biosafety in Microbiological and Biomedical Laboratories (BMBL), U.S. Department of Health and Human Services. It is your responsibility to understand the hazards associated with the material per your organization’s policies and procedures as well as any other applicable regulations as enforced by your local or national agencies.
ATCC highly recommends that appropriate personal protective equipment is always used when handling vials. For cultures that require storage in liquid nitrogen, it is important to note that some vials may leak when submersed in liquid nitrogen and will slowly fill with liquid nitrogen. Upon thawing, the conversion of the liquid nitrogen back to its gas phase may result in the vial exploding or blowing off its cap with dangerous force creating flying debris. Unless necessary, ATCC recommends that these cultures be stored in the vapor phase of liquid nitrogen rather than submersed in liquid nitrogen.
Additional information on this culture is available on the ATCC web site at www.atcc.org.
While every effort is made to insure authenticity and reliability of strains on deposit, ATCC is not liable for damages arising from the misidentification or misrepresentation of cultures.
ATCC recommends that individuals contemplating commercial use of any culture first contact the originating investigator to negotiate an agreement. Third party distribution of this culture is discouraged, since this practice has resulted in the unintentional spreading of contaminated cultures.
1. Vigorously agitate a culture at or near peak density and aseptically transfer a 0.1 ml aliquot to a T-25 tissue culture flask containing 10 ml of fresh complete medium.
2. Screw cap on tightly and incubate at 25°C.
3. Subculture every 10-14d.
2. Aseptically transfer the cell suspension to 15 ml plastic centrifuge tubes.
3. Centrifuge at 200 x g for 5 min.
4. While cells are centrifuging, prepare a 20% solution of DMSO in ATCC Medium 1886.
5. Remove the supernatant and pool the cell pellets into a final volume of 4.5 ml.
6. Combine the cell suspension with an equal volume of 20% DMSO cryoprotectant solution (prepared in step 4) to yield a final concentration of 10% DMSO.
7. Dispense in 0.5 ml aliquots to 1.0-2.0 ml Nunc vials (special plastic vials for cryopreservation).
8. Place the vials in a controlled rate freezing unit. From room temperature cool at -1°C/min to -40°C. At -40°C, plunge ampules into liquid nitrogen.
9. Store ampules in a liquid nitrogen refrigerator until needed.
10. To establish a culture from the frozen state, place a frozen ampule in a 35°C water bath just enough to cover the frozen material. Allow the ampule to thaw completely (2-3 min).
11. Immediately after thawing, aseptically remove the contents and transfer to a T-25 tissue culture flask containing 10 ml of fresh ATCC medium 1886.
12.Screw the cap on tightly and incubate at 25°C. Observe the culture daily. Transfer the culture when many trophozoites are observed.
The product is provided 'AS IS' and the viability of ATCC® products is warranted for 30 days from the date of shipment, provided that the customer has stored and handled the product according to the information included on the product information sheet, website, and Certificate of Analysis. For living cultures, ATCC lists the media formulation and reagents that have been found to be effective for the product. While other unspecified media and reagents may also produce satisfactory results, a change in the ATCC and/or depositor-recommended protocols may affect the recovery, growth, and/or function of the product. If an alternative medium formulation or reagent is used, the ATCC warranty for viability is no longer valid. Except as expressly set forth herein, no other warranties of any kind are provided, express or implied, including, but not limited to, any implied warranties of merchantability, fitness for a particular purpose, manufacture according to cGMP standards, typicality, safety, accuracy, and/or noninfringement.
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This product is sent on the condition that the customer is responsible for and assumes all risk and responsibility in connection with the receipt, handling, storage, disposal, and use of the ATCC product including without limitation taking all appropriate safety and handling precautions to minimize health or environmental risk. As a condition of receiving the material, the customer agrees that any activity undertaken with the ATCC product and any progeny or modifications will be conducted in compliance with all applicable laws, regulations, and guidelines. This product is provided 'AS IS' with no representations or warranties whatsoever except as expressly set forth herein and in no event shall ATCC, its parents, subsidiaries, directors, officers, agents, employees, assigns, successors, and affiliates be liable for indirect, special, incidental, or consequential damages of any kind in connection with or arising out of the customer's use of the product. While reasonable effort is made to ensure authenticity and reliability of materials on deposit, ATCC is not liable for damages arising from the misidentification or misrepresentation of such materials.
Please see the material transfer agreement (MTA) for further details regarding the use of this product. The MTA is available at www.atcc.org.
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David Bushek, personal communication